The identification of small organic molecules that affect specific biological functions is an endeavor that impacts both biology and medicine. Such molecules are useful as therapeutic agents and as probes of biological function. In but one example from the emerging field of chemical genetics, in which small molecules can be used to alter the function of biological molecules to which they bind, these molecules have been effective at elucidating signal transduction pathways by acting as chemical protein knockouts, thereby causing a loss of protein function. (Schreiber et al. J. Am. Chem. Soc. 1990, 112, 5583; Mitchison, Chem. and Biol. 1994, 1, 3) Additionally, due to the interaction of these small molecules with particular biological targets and their ability to affect specific biological functions, they may also serve as candidates for the development of therapeutics.
Because it is difficult to predict which small molecules will interact with a biological target. intense efforts have been directed towards the generation of large numbers, or xe2x80x9clibrariesxe2x80x9d, of small organic compounds. These libraries can then be linked to sensitive screens to identify the active molecules. In many cases, researchers have developed xe2x80x9cbiasedxe2x80x9d libraries, in which all members share a particular characteristic, such as an ability to interact with a particular target ligand, or a characteristic structural feature designed to mimic a particular aspect of a class of natural compounds. For example, a number of libraries have been designed to mimic one or more features of natural peptides. Such peptidomimetic libraries include phthalimido libraries (WO 97/22594), thiophene libraries (WO 97/40034), benzodiazopene libraries (U.S. Pat. No. 5,288,514), libraries formed by the sequential reaction of dienes (WO 96/03424), thiazolidinone libraries, libraries of metathiazanones and their derivatives (U.S. Pat. No. 5,549,974), and azatide libraries (WO 97/35199) (for review of peptidomimetic technologies, see Gante, J., Angew. Chem. Int. Ed. Engl. 1994, 33, 1699-1720 and references cited therein).
Each of these libraries has provided solid phase synthetic strategies for compounds possessing specific core functionalities, but none achieves the complexity of structure found in natural products, or in other lead compounds prepared through traditional chemical synthetic routes. Complex natural products commonly contain several different functionalities and often are rich in stereochemical complexity. Such diversity and complexity are difficult to achieve if the synthesis is restricted to a specific class of compounds.
Recognizing the need for development of synthetic strategies that produce large numbers of complex molecules, Boger et al. (EP 0774 464) have recently developed a solution-phase synthetic strategy for producing a library of compounds based on a functionalizable template core, to which various reagents can be added. However, there remains a need for development of solid-phase strategies, where the more rapid production methods such as split-and-pool strategies can be employed to generate larger ( greater than 1,000,000), more complex libraries. Additional solution-phase strategies would, of course, also be valuable.
The present invention provides methods for the production of compounds and libraries of complex compounds reminiscent of natural products from diversifiable scaffold structures. In particular, the present invention provides synthetic strategies that allow production of complex compounds and preferably large collections of complex compounds that are reminiscent of natural products in that they contain one or more stereocenters, and a high density and diversity of functionality. In preferred embodiments, the compounds of the present inventive libraries are structurally related to a natural product. Alternatively or additionally, the compounds of the inventive libraries possess the capability of acting as a ligand in a biological system to produce a desired inhibitory or promoter effect, and thus may also be functionally reminiscent of natural products.
According to the present invention, the inventive compounds and combinatorial libraries are synthesized from diversifiable solid support bound scaffolds, which are synthesized from readily available or easily synthesizable template structures. In certain embodiments, the inventive compounds and libraries are generated from diversifiable scaffolds synthesized from a shikimic acid based epoxyol template. In other embodiments, the inventive compounds and libraries are generated from diversifiable scaffolds synthesized from the pyridine-based template isonicotinamide.
In addition to providing complex compounds reminiscent of natural products, combinatorial libraries thereof, and methods of their production, the present invention also provides a novel ortho-nitrobenzyl photolinker, and a method for its synthesis, that can be used in the preparation of solid support bound compounds and combinatorial libraries.
The present invention further provides a method for determining one or more biological activities of a library member. In a preferred embodiment, the method for determining one or more biological activities of the inventive compounds comprises contacting the inventive compounds with a biological target, such as a binding target or transcription based assay, and determining a statistically significant change in a biochemical activity relative to the level of biochemical activity in the absence of the compound.
The present invention further provides a kit comprising a library of compounds and reagents for determining one or more biological activities of the library member. To give but one example, the biological activity can be determined by providing a kit containing a binding reagent, such as a direct reagent (binding target) or an indirect reagent (transcription based assay) and a library of compounds.
The present invention additionally provides pharmaceutical compositions containing one or more library members. In a preferred embodiment, the pharmaceutical composition preferably comprises one or more of the inventive compounds and a pharmaceutically acceptable carrier.
xe2x80x9cCombinatorial libraryxe2x80x9d: As used herein, a xe2x80x9ccombinatorial libraryxe2x80x9d is a plurality of complex compounds reminiscent of natural products synthesized from diversifiable scaffold structures by employing different reactants, or monomers, at each stage of the diversification of the scaffold structures. The combinatorial libraries of the present invention may be prepared in solution or on the solid phase.
xe2x80x9cDiversifiable scaffold structuresxe2x80x9d: As used herein, a xe2x80x9cdiversifiable scaffold structurexe2x80x9d is a compound synthesized from a template structure, which contains unique latent or active functionalities capable of being further reacted with synthetic reagents to generate at least one new functionality, but, particularly in the case of a latent functionality, may generate more than one. As used herein, a xe2x80x9clatent functionalityxe2x80x9d is one that is present, but is temporarily inactive. Upon release with an activator or reagent, the latent functionality becomes active, and is thus available for further diversification. For example, a diversifiable scaffold structure may contain an epoxide moiety, which, upon reaction with a nucleophile releases a latent alcohol functionality and generates an additional functionality at the site of nucleophilic attack.
Furthermore, the alcohol functionality can be subsequently diversified using electrophites to yield other functionalities including, but not limited to, ether, ester, carbamate and thioester.
xe2x80x9cComplex compounds reminiscent of natural productsxe2x80x9d: As used herein, a complex compound reminiscent of a natural product is a compound that, similarly to complex natural products which nature has selected through evolution, contains more than one stereocenter, a high density and diversity of functionality, and a diverse range of atoms within one structure. This term can also, for the purposes of the present invention, be used interchangeably with the term xe2x80x9cnatural product-likexe2x80x9d compound. In this context, diversity of functionality can be defined as varying the topology, charge, size, hydrophilicity, hydrophobicity, and reactivity, to name a few, of the functional groups present in the compounds. The term, xe2x80x9chigh density of functionalityxe2x80x9d, as used herein, can preferably be used to define any molecule that contains at least four latent or active diversifiable functional moieties. These structural characteristics may additionally render the inventive compounds functionally reminiscent of complex natural products, in that they may interact specifically with a particular biological receptor, and thus may also be functionally natural product-like.
xe2x80x9cSmall Moleculexe2x80x9d: As used herein, the term xe2x80x9csmall moleculexe2x80x9d refers to an organic compound either synthesized in the laboratory or found in nature. Typically, a small molecule is characterized in that it contains several carbon-carbon bonds, and has a molecular weight of less than 1500, although this characterization is not intended to be limiting for the purposes of the present invention. Examples of xe2x80x9csmall moleculesxe2x80x9d that occur in nature include, but are not limited to, taxol, dynemicin, and rapamycin. Examples of xe2x80x9csmall. moleculesxe2x80x9d that are synthesized in the laboratory include, but are not limited to, the inventive compounds incorporated herein.
xe2x80x9cLinkerxe2x80x9d: The term xe2x80x9clinkerxe2x80x9d, as used herein, refers to a molecule or group of molecules connecting a solid support and a combinatorial library member. The linker may be comprised of a single linking molecule or may comprise a linking molecule and a spacer molecule, intended to separate the linking molecule and the library member by a specific distance.
xe2x80x9cRadially Arrayedxe2x80x9d: The term xe2x80x9cradially arrayedxe2x80x9d as used herein, refers to a spatial arrangement of functionality that projects outwardly in all directions, from the synthesized scaffold structure.
xe2x80x9cProtecting Groupxe2x80x9d: The term xe2x80x9cprotecting groupxe2x80x9d as used herein, refers to a chemical group that reacts selectively with a desired functionality in good yield to give a derivative that is stable to further reactions for which protection is desired, can be selectively removed from the particular functionality that it protects to yield the desired functionality, and is removable in good yield by reagents compatible with the other functional group(s) generated during the reactions.
xe2x80x9cSupportxe2x80x9d: The term xe2x80x9csupportxe2x80x9d, as used herein interchangeably as beads, solid surfaces, substrates, particles, supports, etc. These terms are intended to include 1) solid supports such as beads, pellets, disks, capillaries, pore-glass beads, silica gels, polystyrene beads optionally cross-linked with divinylbenzene, grafted co-poly beads, poly-acrylamide beads, latex beads, dimethylacrylamide beads optionally cross-linked with N,Nxe2x80x2-bis acryloyl ethylene diamine, glass particles coated with a hydrophobic polymer, or any other material having a rigid or semi-rigid surface; and 2) soluble supports such as low molecular weight non-cross-linked polystyrene. These materials also contain functionalities such that identifiers and/or templates, scaffolds, and inventive compounds can be attached to them. It is particularly preferred for the purposes of the present invention that the solid support Tentagel is used.
xe2x80x9cIdentifier Tagxe2x80x9d: The term xe2x80x9cidentifier tagxe2x80x9d as used herein, refers to a means for recording a step in a series of reactions used in the synthesis of a chemical library. For the purposes of this application, the terms encoded chemical library and tagged chemical library both refer to libraries containing a means for recording each step in the reaction sequence for the synthesis of the chemical library.